1. Field of the Invention
This invention relates to a novel gene detection method to detect a certain gene specifically and a device for such detection.
2. Description of the Related Art
A genetic information stored in DNA is expressed as a protein or an enzyme through mRNA. By the effects of such protein or enzyme, various compounds necessary to maintain the vital actions are biosynthesized and metabolized. Thus a life is present as a dynamic equilibrium system of various substances controlled by genes.
There are 50 to 100 thousands of human genes. When some of them involve abnormality or change, such as defect or duplication, the characteristics, types and amounts of the proteins synthesized are changed, resulting in the poorly balanced biosystem, which may cause diseases. Thus, by detecting known pathogenic genes, the diseases may be identified or prevented. Such diagnosis based on the genes themselves has been developed as a results of the recently advancing technology of gene engineering, and is called as gene diagnosis.
When compared with conventional diagnostic methods, the gene diagnosis has characteristics as mentioned below.
Considering the mechanisms of gene expression, it can be presumed that changes in genes occur prior to almost all biochemical changes. Therefore, gene diagnosis by means of detecting the genetic change enables the diagnosis or prognosis prior to development of a disease which is one of phenotypes. Accordingly, the diagnosis and prognosis can be conducted before the development, in the latent period or at the earliest stage of the disease. This is the primary characteristic. As the secondary characteristic, gene diagnosis relating to the genetic diseases is independent from the organs or tissues to be analyzed since all genes in a living body are the same. This is particularly important in the diagnosis in fetus. Thus, this secondary characteristic enables the diagnosis simply by sampling amniotic fluid from a pregnant woman and analyzing the fetal cells suspending in the amniotic fluid.
Procedure of gene diagnosis conventionally employed is summarized as follows.
Genes are extracted from a samples and cleaved, if necessary, by appropriate restriction enzymes, and then subjected to electrophoresis and southern blotting. Then a nucleic acid probe (usually radiolabelled) having the base sequence complementary to the gene to be detected is hybridized to the blotted gene. Subsequently, the hybridized nucleic acid probe is detected by exposing an X-ray film to the radiation emitted from the labeled probe at lower temperature to confirm the presence of the gene.
The conventional detection method mentioned above involves the limitation of the place of diagnosis due to the use of radioisotopes and should be conducted with sufficient care of handling reagents. For the purpose of reducing such inconvenience, safe labeling agents substituting the radioisotopes are being developed and several detection methods, in which probes are utilized, such as avidin-biotin bond method or enzymatic or fluorescent method and the like have already been suggested. However, these methods can not achieve the sensitivity superior to that of the method using radioisotopes. They also involve the problems of the time period required to detecting the gene as long as 2 or 3 days as well as complicated procedure of determination.
On the other hand, quantification of a certain antigen or antibody present in a sample generally employs radioimmunoassay (RIA). However, RIA requires special instruments and authorized operators therefor capable of handling radioisotopes since this method also employs radioisotopes similarly as in the gene diagnosis methods mentioned above. In addition, waste disposal in this method should be done with particular care. As one of the other analytic methods, immunoelectrophoresis, which requires a long period for determination and has a poor sensitivity, can be suggested, although this method is not applicable in case of the samples containing only trace amount of test substance.